Selectin E/CD62e ELISA Kit

Catalog No. ABIN1112589

Product Details Selectin E/CD62e ELISA Kit

Target: Selectin E/CD62e (SELE) (Selectin E (SELE))

Reactivity: Mouse

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Detection Range: 500-10000 pg/mL

Minimum Detection Limit: 500 pg/mL

Application: ELISA

Purpose: For quantitative detection of E-Selectin in mouse serum, plasma, urine, cell culture supernatant or tissue samples.

Sample Type: Serum, Plasma, Urine, Tissue Samples, Cell Culture Supernatant

Analytical Method: Quantitative

Components: 1. One 96-well plate pre-coated with anti-mouse E-Selectin antibody 2. Standard: 0.5ml (13,500pg /mL) 3. Standard diluent buffer: 1.5 ml 4. Wash buffer (30x): 20 ml.

Material not included: 1. 37 °C incubator 2. Microplate reader (wavelength: 450nm) 3. Precise pipette and disposable pipette tips 4. Automated plate washer 5. ELISA shaker 6. 1.5ml of Eppendorf tubes 7. Plate cover 8. Absorbent filter papers 9. Plastic or glass container with volume of above 1L

Application Details

Comment:

This kit was based on standard sandwich enzyme-linked immune-sorbent assay technology. The purified anti-E-Selectinantibody was pre-coated onto 96-well plates. And the HRP conjugated anti-E-Selectin antibody was used as detection antibodies. The standards test samples and HRP conjugated detection antibody were added to the wells subsequently mixed and incubated then unbound conjugates were washed away with wash buffer. TMB substrates (A & B) were used to visualize HRP enzymatic reaction. TMB was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the E-Selectin amount of sample captured in plate. Read the O.D. absorbance at 450nm in a microplate reader and then the concentration of E-Selectin can be calculated.

Plate: Pre-coated

Reagent Preparation:

1. Before the experiment, centrifuge each kit component for several minutes to bring down all reagents to the bottom of tubes. 2. It is recommend to measure each standard and sample in duplicate. 3. Do NOT let the plate completely dry at any time! Since the dry condition can inactivate the biological material on the plate. 4. Do not reuse pipette tips and tubes to avoid cross contamination. 5. Do not use the expired components and the components from different batches. 6. To avoid the marginal effect of plate incubation for temperature differences (the marginal wells always get stronger reaction), it is recommend to equilibrate the ABC working solution and TMB substrate for at least 30 min at room temperature (37°C ) before adding to wells.The TMB substrate (Kit Component 8) is colorless and transparent before use, if not, please contact us for replacement.

Sample Preparation:

Preparation of sample and reagents 1. Sample Isolate the test samples soon after collecting, then, analyze immediately (within 2 hours). Or aliquot and store at -20 °C for long term. Avoid multiple freeze-thaw cycles. Serum: Coagulate at room temperature for 10-20 °C min, then, centrifuge at the speed of 2000-3000 r.p.m. for 20 min to collect supernatant. If precipitation appeared, centrifuge again. Plasma: Collect plasma using EDTAor citrate plasma as an anticoagulant, and mix for 10-20 °C min, centrifuge at the speed of 2000-3000 r.p.m. for 20 min of collection. If precipitation appeared, centrifuge again. Urine: Collect urine using a sterile container, centrifuge at the speed of 2000-3000 r.p.m. for 20 min to collect supernatant. If precipitation appeared, centrifuge again. For collection of hydrothorax and cerebrospinal fluid, take reference to this operation. Cell culture supernatant: For secretory components: use a sterile container to collect. Centrifuge at the speed of 2000-3000 r.p.m. for 20 min to collect supernatant. For intracellular components: Dilute cell suspension with PBS(pH7.2-7.4) to make the cell concentration reached 1 million / ml. Damage cells and release of intracellular components through repeated freeze-thaw cycles. Centrifuge at the speed of 2000-3000 r.p.m. For 20 min to collect supernatant. If precipitation appeared, centrifuge again. Tissue samples: Cut samples and weight, add certain volume of PBS (pH7.4), rapidly frozen with liquid nitrogen. After melting, store samples at 2-8℃ . Add certain volume of PBS (pH7.4), homogenize thoroughly, centrifuge at the speed of 2000-3000 r.p.m. for 20 min to collect supernatant. Note: 1. Coagulate blood samples completely, then, centrifuge, and avoid hemolysis and particle. 2. NaN 3 can not be used as test sample preservative, since it is the inhibitor for HRP. 3. After collecting samples, analyze immediately or aliquot and store frozen at -20 °C. Avoid repeated freeze-thaw cycles. 2. Wash buffer Dilute concentrated Wash buffer (Kit Component 4) 30-fold (1:30) with distilled water (i.e. add 20 ml of concentrated wash buffer into 580 ml of distilled water). 3. Standard Reconstitution of the Lyophilized Mouse E-Selectin standard (Kit Component 2): standard solution should be prepared no more than 2 hours prior to the experiment. Two tubes of standard are included in each kit. Use one tube for each experiment. (Note: Do not dilute the standard directly in the plate) a. 10,000 pg/ml of standard solution: Add 0.5 ml of the 13,500pg/ml standard (Kit Component 2) into 0.175 ml Standard diluent buffer (Kit Component 3) and mix thoroughly. b. 5,000 pg/ml -> 625 pg/ml of standard solutions: Label 4 Eppendorf tubes with 5,000 pg/ml, 2500pg/ml, 1250 pg/ml, 625 pg/ml, respectively. Aliquot 0.2 ml of the Standard diluent buffer (Kit Component 3) into each tube. Add 0.2 ml of the above 10,000 pg/ml standard solution into 1st tube and mix thoroughly. Transfer 0.2 ml from 1st tube to 2nd tube and mix thoroughly. Transfer 0.2 ml from 2nd tube to 3rd tube and mix thoroughly, and so on. 5000 2500 1250 625 0.2ml 0.2 ml 0.2 ml 0.2 ml E-Selectin Protein Standards [pg/ml] 100 pg/ml m. E-Selectin Solution 10,000 Chongqing Biospes Co., Ltd Product Manual

Restrictions: For Research Use only

Handling

Preservative: Sodium azide, Thimerosal (Merthiolate)

Target Details for Selectin E/CD62e

Target: Selectin E/CD62e (SELE) (Selectin E (SELE))

Alternative Name: E-Selectin (SELE Products)

Synonyms: SELE ELISA Kit, CD62E ELISA Kit, ELAM ELISA Kit, ELAM1 ELISA Kit, ESEL ELISA Kit, LECAM2 ELISA Kit, E-selectin ELISA Kit, Elam ELISA Kit, selectin E ELISA Kit, selectin, endothelial cell ELISA Kit, SELE ELISA Kit, Sele ELISA Kit

Background: E-selectin, also known as CD62 antigen-like family member E (CD62E), endothelial-leukocyte adhesion molecule 1 (ELAM-1), or leukocyte-endothelial cell adhesion molecule 2 (LECAM2), is a cell adhesion molecule expressed only on endothelial cells activated by cytokines. The ELAM gene is present in single copy in the human genome and contains 14 exons spanning about 13 kb of DNA . E-selectin mediates the adhesion of tumor cells to endothelial cells, by binding to E-selectin ligands expressed by neutrophils, monocytes, eosinophils, memory-effector T-like lymphocytes, natural killer cells or cancer cells.

Pathways: Thromboxane A2 Receptor Signaling